Buoy suspension fractionation system

ABSTRACT

A separator that uses centrifugation to fractionate a suspension such as blood comprises a separation container and a buoy. The buoy is carried in the separation container and has a tuned density that is configured to reach an equilibrium position in a suspension. The guide surface is carried on the buoy upper surface and is inclined to an accumulation position near a buoy perimeter. The buoy suspension fractionation system can be used in a method of isolating a fraction from a suspension, and in a method for re-suspending particulates for withdrawal.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application includes subject matter similar to concurrently filedU.S. patent application Ser. No. 12/101,594, filed on Apr. 11, 2008, nowU.S. Pat. No. 7,806,276, issued on Oct. 5, 2010, incorporated herein byreference.

This application claims the benefit of U.S. Provisional Application No.60/911,407, filed on Apr. 12, 2007. The disclosure of the aboveapplication is incorporated herein by reference.

FIELD

The present teachings relate to a separator that uses densitydifferences to fractionate a suspension such as blood.

BACKGROUND

Clinicians have identified a wide range of therapeutic and laboratoryapplications for autologous isolated fractions, such as plateletconcentrate, platelet-poor-plasma, and stromal cells, of suspensionssuch as blood, bone marrow aspirate, and adipose tissue. Cliniciansgenerally prefer to draw and fractionate the autologous suspension atthe point-of-care. Point-of-care fractionation can reduce the need formultiple appointments to draw and fractionate the autologous suspensionwhich can be costly and inconvenient. Additionally, point-of-carepreparation reduces potential degradation of the autologous suspensionthat can begin once the autologous suspension is removed from a patient.Point-of-care fractionation systems should be easy to operate to reducethe need to provide clinicians with extensive instruction, quick so thetherapeutic fraction can be isolated and administered during a singlepatient visit, efficient to effectively isolate the fraction to adesired concentration, and reproducible to operate over wide variationsin suspension characteristics. An example of a buoy based suspensionfractionation system is shown in Biomet Biologics, Inc. internationalbrochure entitled “Gravitational Platelet Separation System Acceleratingthe Body's Natural Healing Process.” 2006

SUMMARY

A buoy suspension fractionation system comprises a separation containerand a buoy. The separation container defines a volume enclosed by acontainer wall, a container bottom, a container top and an access portto access the volume. The buoy is carried in the separation containerand has a tuned density that is configured to reach an equilibriumposition in a suspension. The buoy comprises a buoy upper surface and abuoy sidewall defining a height, a transverse dimension, and aperimeter. The buoy further comprises a guide surface and a collectionspace above the buoy upper surface. The guide surface is carried on thebuoy upper surface and is inclined to an accumulation position near thebuoy perimeter. The buoy suspension fractionation system can be used ina method of isolating a fraction from a suspension, and in a method forisolating a fraction and re-suspending isolated particulates forwithdrawal.

BRIEF DESCRIPTION OF THE DRAWINGS

The present teachings will become more fully understood from thedetailed description and the accompanying drawings, wherein:

FIG. 1 is an environmental view of a fractionation device including asuspension fractionated during the centrifuge process;

FIG. 2 is an environmental view of a suspension being added to afractionation device;

FIG. 3 is an environmental view of a centrifuge;

FIG. 4 is an environmental view of a first fraction being removed fromthe fractionation device;

FIG. 5. is an environmental view of the fractionation device beingagitated to re-suspend a portion in a second fraction;

FIG. 6 is an environmental view of the second fraction being removedfrom the fractionation device;

FIG. 7 is an environmental view of a therapeutic application of thesecond fraction;

FIG. 8 is an environmental view of a separation container and a buoy;

FIG. 9A is a plan view of a buoy according to various embodiments;

FIG. 9A1 is a plan view of a buoy at a selected transverse plane;

FIG. 9A2 is a plan view of a buoy at a selected transverse plane;

FIG. 9B is a cross-sectional view of the buoy of FIG. 2A;

FIG. 10 is a perspective view of a buoy, according to variousembodiments;

FIG. 11A is a perspective view of a buoy, according to variousembodiments;

FIG. 11B is a perspective view of a buoy in a closed position, accordingto various embodiments;

FIG. 12 is a perspective view of a buoy, according to variousembodiments;

FIG. 13 is a perspective view of a buoy, according to variousembodiments;

FIG. 14 is a plan view of a buoy, according to various embodiments;

FIG. 15 is a plan view of a buoy, according to various embodiments;

FIG. 16. is a plan view of a buoy, according to various embodiments;

FIG. 17 is a plan view of a buoy, according to various embodiments;

FIG. 18 is a plan view of a buoy, according to various embodiments;

FIG. 19 is an environmental view of a selected component being withdrawnfrom a separation device according to various embodiments; and

FIG. 20 is a kit according to various embodiments, for separation andextraction of a selected component of a suspension.

DETAILED DESCRIPTION OF VARIOUS EMBODIMENTS

FIG. 1 shows a buoy suspension fractionation system 10, according tovarious embodiments that can be used in a clinical or laboratoryenvironment to isolate fractions from a suspension or multi-componentmaterial removed from a patient or a preparation of extracted or excisedmaterial from a patient. The suspension can include a sample of blood,bone marrow aspirate, cerebrospinal fluid, adipose tissue, and theisolated fractions can include platelets, platelet poor plasma, plateletrich plasma and stromal cells. The isolated fractions can each haveequilibrium point or positions within the separation container that areachieved when separation has occurred. For example, a buffy coat ofwhole blood may have an equilibrium position above that of the red bloodcells when a sample of whole blood is separated.

Isolated fractions can be used in a variety of clinical applications,animal applications, and laboratory applications. Some of the clinicalapplications include peripheral vascular disease, orthopedic surgery,plastic surgery, oral surgery, cardio-thoracic surgery, brain and neuralprocedures, and wound healing. Laboratory applications includeisolating, creating or synthesizing therapeutic materials or materialsfor analysis from fractions produced by the fractionation system.

Although the fractionation system 10 can be used allogeneically, such aswith pooled blood, the fractionation system 10 can be used autologouslyto reduce risks of potential incompatibility and contamination withpathogenic diseases. Also, other autologous materials can be usedincluding cerebrospinal fluid, cerebrospinal fluid can be obtained via aspinal tap or other appropriate collection procedure. A generaldescription of a fractionation system is provided in a Biomet Biologics,Inc. international brochure “Gravitation Platelet Separation SystemAccelerating the Body's Natural Healing Process” (2006) and adescription of a therapeutic procedure using platelet concentrate isshown in a Biomet Biologics, Inc. international brochure “ShoulderRecovery with the GPSO® Platelet Concentration System” (2004),incorporated herein by reference.

FIGS. 2-7 show exemplary fractionation system operational steps for aclinical therapeutic application embodiment. The operational steps beginin FIG. 2 by inputting autologous (although pooled blood can be used)whole blood into the fractionation system 10, via an access port 22. Thefractionation system 10 is placed into a centrifuge 23 in FIG. 3 andspun about five minutes to about twenty minutes at a rate of about 320rpm to about 5000 rpm (this speed may produce a selected gravity thatmay be approximately 7.17×g to about 1750×g (times greater than thenormal force of gravity)). The first fraction or top fraction 308 (FIG.1), which can be platelet-poor-plasma according to various embodimentsincluding from a whole blood sample, is shown being removed in FIG. 4.The fractionation system 10 is agitated in FIG. 5 to re-suspend at leasta portion of a second fraction 310, which can be platelet-rich-plasma orplatelet concentrate, according to various embodiments including fromwhole blood fractionation. The second fraction is removed from thefractionation system 10 in FIG. 6. Finally, the second fraction isapplied as part of a therapy, such as shown in FIG. 7 to treat elbowtendonitis. The second fraction can be injected into a selected portionof an elbow 29 to treat tendonitis.

It will be understood that the buoy 30 can be altered depending upon thematerial placed in the container 12. For example, if neural stem cellsare to be separated from cerebrospinal fluid then the buoy 30 can have adensity to allow collection of the neural stem cells in the collectionarea 52 of the system 12. The collected neural stem cells can also beapplied for therapeutic reasons or used in laboratory study, isolation,culture, etc.

Returning reference to FIG. 1 and with additional reference to FIGS.8-9B, the suspension fractionation system 10 comprises a separationcontainer 12 and a buoy 30. The separation container 12 can be aseparation tube having a container wall 16, a container bottom 18, and acontainer top 20 enclosing a volume 21 that can be accessed by one ormore access ports 22, 26, 27, and a container vent 31. The container 12may be formed of any appropriate material, such as the Cryolite Med® 2material sold by Cyro Industries Evonik Degussa Corp. The container 12can be about 50 mm to about 150 mm in height, including about 102 mm inheight. The container 12 can have an internal diameter of about 20 mm toabout 40 mm, including about 25 mm to about 35 mm and define a volume ofabout 30 ml to about 100 ml, including about 30 ml to about 60 ml. Theseparation container 12 can have any appropriate shape, such as an oval,provided the buoy 30 is shaped to conform to the separation container12. Though not particularly illustrated, the separation container 12 canalso have more than one compartment, such as a separation tube and anarea to transfer tube contents away from the separation tube 12. Forexample, a separate compartment can be formed to house the assembly ofthe buoy 30 and isolator 32 separate from another area.

The various ports 22, 26 and 27 can be provided to allow access to anyappropriate compartment of the container 12. The access ports 22, 26, 27can be any means that allow communication from outside the separationcontainer 12 to the separation container volume 21 such as a Luer lockport, a septum, a valve, or other opening. The container vent 31 allowsmovement of air between the inside and outside the separation container12 to equalize pressure when suspension in introduced into or withdrawnfrom the separation container 12. The container vent 31 can include avent filter 31 a to serve as a sterile barrier to allow air to enter theseparation container 12 while preventing undesired materials fromentering the separation container 12.

When the separation container 12 is at rest, a buoy perimeter 30 a andthe container wall 16 can be dimensioned to form an interference fit tohold the buoy 30 at a position in the separation container 12. When theseparation container 12 is centrifuged, the buoy perimeter 30 a and thecontainer wall 16 have clearance allowing the buoy 30 to move within theseparation container 12 and a material to pass between the buoyperimeter 30 a and the container wall 16. For example, the container 12can compress axially to increase its internal diameter. Alternatively,the buoy 30 could have an opening (e.g. FIG. 16), such as a centrally orinternally located opening 176 or a peripheral channel 168 a (FIG. 13)running the height of the buoy, which would allow a material to movethrough the buoy.

The buoy 30 is carried in the separation container 12 and has a tuneddensity that is configured to reach a selected equilibrium position in asuspension. The buoy can have its density tuned in the range from about1.0 g/cc to about 1.10 g/cc, such as about 1.06 g/cc. The buoy 30,according to various embodiments, can be formed to include the tuneddensity and can be formed of one or more materials to achieve the tuneddensity.

For example, the density of about 1.06 g/cc can position the buoy 30, ora selected part of the buoy 30 including the collection area 52, at anequilibrium position of a buffy coat of a separated whole blood sample.In a further example, the density can also be tuned so that thecollection area 52 is near an equilibrium position, such as where neuralstem cells collect in a selected suspension. Regardless of the densityof the buoy 30, it can be selected to position the buoy 30 at anequilibrium position of a selected material.

As illustrated in FIG. 1, the collection area 52 is positioned withinthe container 12 after a separation procedure has occurred. Thecollection area, defined relative to the buoy 30, is positioned at theequilibrium position of the separated or isolated fraction 310 in thecontainer. The equilibrium position of a selected fraction can bedefined as its position within the container relative to other fractionsin the container of a separated sample or material. The equilibriumposition can also be defined relative to the axis X of the buoy 30 orthe container 12. The equilibrium position, however, may depend upon theamount of the sample of the amount of a selected fraction within asample. According to the illustration in FIG. 1, the equilibriumposition of the fraction 308 is above or nearer the top 20 of thecontainer 12 than the equilibrium position of the fraction 310. Thus,the buoy 30 can be tuned, such as including a selected density orspecific gravity, to position the collection area 52 relative to anequilibrium position of any selected fraction.

The buoy comprises a buoy upper surface 48 and a buoy sidewall 38, 40defining a height H1, H2, a transverse dimension at planes A₁, A₂, and aperimeter 30 a, discussed further herein. The buoy further comprises aguide surface 42. In some embodiments, the buoy can further comprise acollection port 50 and a precision collection region 44. The collectionport 50 communicates with the access port 27 and communicates with acollection space 52 above the buoy upper surface 42 and can be locatednear the buoy perimeter 30 a. In some embodiments, the collection port50 is not carried on the buoy, but rather the collection port is awithdraw device such as a syringe that is inserted through an accessport or top of the tube 12.

With reference to FIG. 9A, the buoy 30 has a first height dimension H1,a second height dimension H2, a maximum width or transverse crosssectional area W1 at plane A1, a second width or transverse crosssectional area W2 at plane A2, a guide surface angle ∝, and precisioncollection area 44 including a surface 46 defining a precisioncollection region angle β. The height of the buoy 30, according tovarious embodiments, can be defined relative to a central axis X, whichcan also be a longitudinal axis X of the container 12. The sidewalls ofthe buoy 30 and the container 12 can also be substantially parallel tothe axis X. Although certain dimensions are shown in FIG. 9A, the buoyperimeter could be shaped differently provided the perimeter conforms tothe separation container 12.

The guide surface 42 is carried on and/or defined by the buoy uppersurface 48 and is inclined to an accumulation position at or near thebuoy perimeter. The guide surface 42 serves as a guide means forconveying particles down an incline toward an equilibrium interface orcollection region. The guide surface 42 can be inclined relative to thebuoy sidewall 38 height for a distance of more than one-half the buoytransverse dimension or width W1, such as about two-thirds the buoytransverse dimension, and in various embodiments the guide surface canbe inclined relative to the buoy sidewall 38 substantially throughout alength of the guide surface 42.

The guide surface 42 can be substantially planar and can have an averageangle in the range from the minimum for particulates to move down theguide surface, regarding blood platelets, for example, about 10 degreesto about 60 degrees. For example, angle α can be about 5 degrees toabout 89 degrees, or greater, including about 30 degrees to about 89degrees. Angle α can, exemplary, be exactly or about 60 degrees invarious embodiments. In some embodiments, the guide surface can includecontours defined in the guide surface with multiple angles such as shownin FIGS. 10 and 12. For example, in FIG. 10, a buoy 80, according tovarious embodiments, can include two guide surface contour walls 96, 98to assist in defining a guide surface 100. The two walls 96, 98 candefine a trough that extends a selected distance across the guidesurface 100, such as more than two thirds. The trough can define an areaof the guide surface that is lower than the surrounding area. Acontoured precision collection region 92 can also be defined thatcommunicates with a port 94. In FIG. 12, a buoy 140 can include a guidesurface 152 that includes two inclined sides 154, 156 angled towards aselected region, such as a center of the guide surface 152. The entireguide surface can also be inclined towards a collection port 158, in anamount as discussed above.

In various embodiments, as exemplary illustrated in FIGS. 9A, 9A1, and9A2 the different buoy transverse cross-sectional areas W1, W2 can bedefined at various planes, such as A₁, A₂, etc. As illustrated, varioustransverse cross-sectional areas can be defined by the buoy 30 due tothe angled top wall 42. The transverse cross-sectional areas defined atthe various planes A₁, A₂ can be positioned at selected locations basedupon characteristics of the buoy 30, such as density. The height H2,angle α, etc. The width dimension can be 1 inch to about 2 inchesincluding about 1.347 inches (about 25 mm to about 51 mm, includingabout 34.21 mm) for W2. The dimension of W1 can depend upon the selectedlocation of plane A1. These dimensions can achieve various areasdepending upon the geometry of the buoy 30. Nevertheless, the area atplane A2 can be substantially similar to an area at a transverse planewithin the container 12.

In use, the substantially maximum transverse cross-sectional area W1 ofthe buoy 30 can be positioned at a selected location. As illustrated inFIG. 9A1, the maximum cross-sectional area is at plane A₁. The plane A₁can be positioned at or near a selected equilibrium interface, in use.The position of the plane A₁ is selected by selecting a density of thebuoy 30 and the known or estimated density of the material into whichthe buoy 30 is positioned. The buoy's maximum transverse cross-sectionalarea near the intended or selected interface results in a substantiallymaximum change in displacement of the relative volume of a fractionbelow the equilibrium interface and substantially maximum change indisplacement of a fraction above the equilibrium interface relative tochange in the axial orientation of the buoy relative to the interface.This can improve fractionation isolation by ensuring that the maximumtransverse cross-section displaces a maximum amount of area within thecontainer 12 at the selected interface. For example, more than 90% of awhole blood's platelets can be isolated.

Thus, in applications involving suspensions, such as whole blood, whichmay be variable in composition between samples, sample density variationwill result in minimal variation in the axial orientation of the buoyrelative to a selected equilibrium interface. The minimal variation inaxial location of the buoy 30 in the container 12 is based at least inpart on the maximum displacement of a material in the container at themaximum transverse cross-section of the buoy 30. In other words, foreach small variation of axial location of the buoy 30, a maximumdisplacement occurs. In selected uses, the buoy's maximumcross-sectional plane A₁ is provided at a selected location and theminimal axial variation helps to ensure the plane A₁ is properly placed.

Additionally, at or near the buoy's maximum transverse cross-sectionalarea, the cross-sectional area of the fractionated material is nearminimal. Simply, within the container 12 at a selected position if amaximum transverse cross-section of the buoy 30 is at a selectedposition, then a relatively minimal amount of other material can bepresent at the same location. In combination, the minimization ofcross-sectional area of fractionated material and minimization ofvariation of axial orientation of the buoy in relation to an equilibriuminterface results in minimization of variability of fractionatedmaterial volume near the interface.

The precision collection region 44, 92 (FIGS. 9A, 9B, and 10) can beinterposed between the guide surface and the accumulation position at ornear the buoy perimeter. The precision collection region 44, 92 servesas a precision collection structure for collecting a precise, high yieldand/or pure amount of a selected fraction. The precision collectionregion 44, 92 can be raised or lowered in relation to the buoy perimeterto vary the fraction in the collection region without the need to makesubstantial changes to other buoy design features. In other words, thedimension H1 can be changed. Generally, the height H2 can be about 2.5mm to about 5.1 mm. The height H1 will generally be constrained by theheight H2 and the angle α. According to various embodiments, theprecision collection region 44 is shown in FIG. 9A formed at an angle βin relation to the sidewall 40. The angle β can be any appropriate anglesuch as about 10 degrees to about 60 degrees, including about 45degrees. According to various embodiments, the precisions collectionregion 92 can be contoured, FIG. 10.

According to various embodiments, an isolator 32, is coupled to the buoy30. The combination of the isolator and buoy, according to variousembodiments, can also be referred to as a separation assembly member.Exemplary isolators 82, 122, 170, 180, 190 are illustrated coupled toexemplary buoys 80, 120, 140, 160, 182, 192. The isolator 32, forexample, provides a means for creating the collection compartment 52 andcomprises one or more spacers 58, 60 to position the isolator 32 apartfrom the buoy 30 to create the collection compartment 52. A withdrawport 70 can be carried on the isolator 32 communicating with thewithdraw port 27 and the collection port 50. The spacer 58, 60 can alsoserve as a conduit 68 between the collection port 50 and a withdraw orwithdraw port 27. The withdraw port 27 serves as a structure forwithdrawing the isolated or second fraction 310 from the collectioncompartment 52.

The isolator 32 can be configured from a material with a lower densitythan the buoy 30, such as a density of about 1.0 g/cc or less. A volumeof the isolator 32 can be substantially less than a volume of the buoy30. The isolator 32 can be configured so the isolator volume and thebuoy volume combined below a selected equilibrium interface are greaterthan the isolator volume and the buoy volume combined above theequilibrium interface. As discussed above, an equilibrium interface caninclude a position relative to the platelet concentrate or buffy coatfrom a centrifuged whole blood sample, such as at or just below theplatelet concentrate or buffy coat. By configuring the isolator 32 andbuoy 30 with more volume below the equilibrium interface than above theequilibrium interface, the buoy 30 operates in a more repeatable mannereven between a wide range in variations in compositions such as wholeblood where the variability in density of a more dense fraction (e.g.red blood cells) is less than the variability in density of a less densefraction (e.g. plasma). For example, the make up of a whole blood samplefrom one patient to the next can be markedly different.

Between individual patients, the density of the red blood cell orerythrocyte fraction of a whole blood sample can generally vary lessthan the density of a plasma or serum portion of a whole blood sample.Therefore, positioning a greater volume of the isolator and buoy withinthe denser fraction can assist in having highly repeatable and highlyefficient collection or separation of a whole blood sample. The heightH2 can be varied or selected to ensure a maximum or selected volume ofthe isolator and buoy are positioned within the denser fraction of thewhole blood sample.

According to various embodiments, the isolator may include variousfeatures. An isolator 122 can be configured to move relative to a buoy120, as illustrated in FIGS. 11A and 11B. The isolator 122 can movealong a column or spacer 132 in the direction of arrow 123 duringextraction of a selected fraction. The isolator 32, 82 can also besubstantially uniformly thick or vary in thickness 122, 180, 190.

An isolator 170 can include collection openings 174 (FIG. 13). Theisolator 32 can also include a collection vent 67 (FIG. 9A), which canalso include a collection valve, a collection passage, or a collectionvent tube or passage 203. The collection openings 174 can reduce thedistance particles, such as platelets which are fragile and adherent,travel to reach a guide surface 162 and reduce the time that particlesare in contact with surfaces. Various types of collection openings canbe used.

The collection openings 174 can be sized to permit selected particles topass yet sufficiently small so suspension fluid tension maintainsadequate isolation of the collection compartment. The collectionopenings can also include various valves such as a duck bill or flapperbill which can open under certain conditions and close under others. Acollection valve can be interconnected with any appropriate portion suchas with a collection port 70 or passage 68.

The collection vent passage 67 through the isolator 32 equalizespressure when fluid is withdrawn from the collection area 52. The spacer58 can serve as a conduit for the collection vent passage 67, thecollection port 50, or both. The collection valve communicates with thecollection vent passage 67 to control collection vent passage 67operation and close the collection vent passage 67 during re-suspensionagitation. The collection vent tube 203 communicates with the collectionvent passage 67 and air. The air can be the air above the collectionarea 52 (i.e. a portion of the suspension above the isolator 32 has beenremoved) or through an opening 205 in the container wall and generallythrough a sterile barrier (e.g. a sterile foam filter). The collectionvent tube 203 allows removal of fractionated suspension in thecollection compartment without the need to remove the fraction, such asplasma, above the isolator 32. Although, without a collection vent tube203, the fraction above the isolator could be removed and the collectionarea could be vented to the area above the isolator.

Various embodiments further comprise a mechanical agitator 130 carriedin a collection compartment 128 (for example FIGS. 11A and 11B).

The isolator 122 is moveable relative to the buoy 120. The isolator 122can be in an open position after centrifugation of the separationcontainer. During removal of material from the collection compartmentthrough the collection port 134, the isolator 122 can move in thedirection indicated by arrow 123 toward the buoy 120 to decrease orclose the volume of the collection compartment 128.

The buoy 30 can also be formed in a plurality of selectable sizes,having different dimensions, such as those illustrated in FIG. 9A. Theaxial dimensions of the buoy 30 can be selected to achieve anappropriate displacement of the suspension in the container 12,especially after fractionation has occurred. Angle α, defined betweenthe outer edge 38 and the surface 42 can be any selected angle. Forexample, angle α can be about 30 degrees to about 89 degrees, includingabout 60 degrees. The angle α can generally be created to be as small aspossible to allow a steep angle of the surface 42 towards the inlet port50 that will not damage the material being collected within thecollection space 52. As discussed above, the height H2 can be selectedto determine or select the amount of the buoy 30 positioned within aselected fraction, such as a dense fraction, of a sample separatedwithin the separation system. Height H2 can be about 0.1 inches to about0.2 inches, including about 0.18 inches (about 2.5 mm to about 5.1 mm,including about 4.57 mm). An exemplary height H2 is 0.1795 inches (4.559mm), depending upon selected applications, the size of the separationsystem, and other selected factors. Nevertheless, the height H1 isgenerally defined by the height H2 and the angle α. Height H1 can beabout 0.8 inches to about 1.2 inches, including about 1 inch (about 20mm to about 30 mm, including about 25 mm). An exemplary height H1 caninclude 1.0 inches (25 mm). The positioning of the collection area 52,including the inlet port 50, can be based upon the height H2 and how thebuoy 30 interacts with the material into which it is positioned, via theheight H2.

A buoy 182, as illustrated in FIG. 17, can include an isolator 180positioned relative thereto. The isolator 180 can include a centerplaced substantially over a center of the buoy 182. The center of thebuoy 182 and the isolator 182 can both be defined by peaks or apexes 184and 186, respectively.

A buoy 192, as illustrated in FIG. 18, can also be positioned relativeto an isolator 190. The buoy 192 can include an apex 194 near a centerof the buoy 192 and the guide surface extending from an edge of the buoy192 to a second edge of the buoy 192. The isolator 190 can also includean apex 196 generally near its center. The isolator 190 can also includea surface 198 that extends from one edge of the isolator to another edgeof the isolator 190.

The isolators 180, 190 can act substantially similar to the isolator 32,discussed above. The isolator 180, 190 can define an angle between anapex or the withdrawal port 70 and an outer edge of the isolators 180,190. The upper surface of the isolators can include an angle to assistin directing a selected material, such as a platelet fraction of wholeblood sample, to the collection area or surface 42 of the buoys 182,192. Generally, the isolators 180, 190 can include a height or volume tosubstantially minimize the volume of the isolator 180, 190 relative tothe buoys 182, 192. As discussed above, this can assist in positioningthe buoys 182, 192 relative to a dense (e.g. red blood cell) fraction ofa whole blood sample. The angle of the isolators 180, 190 and the heightof the isolators 180, 190 can be selected to provide for a minimaldistance of travel or least disturbance of a selected collected fractionof a material, such as a whole blood sample.

As discussed above, the buoy suspension fractionation system 10 can beused in a method of isolating a fraction from a suspension. Theseparation container 12 can be centrifuged for a period that isappropriate for the suspension. The buoy 30 in the separation container12 is allowed to reach an equilibrium position within the formedfractions. Typically, the buoy moves from the separation containerbottom to an equilibrium position within and/or between the fractions.In some embodiments, the buoy 30 is configured with the transversedimension cross-sectional area of the buoy near the equilibriuminterface to be substantially the buoy's maximum transversecross-sectional area A₁, as illustrated in FIG. 1. As discussed above,the design of the buoy can be determined to position a maximum crosssectional area of the buoy within a selected fraction, such as the redblood cell fraction, of a whole blood sample. The positioning of thebuoy can be based upon the density of the buoy which is determined fromthe density of a selected fraction, such as a red blood cell fraction.Therefore, the buoy can be created or formed to include a density tosubstantially position it within a red blood cell fraction, for example,of a sample to be separated. For example, the buoy can have a density ofabout 1.010 g/cc to about 1.1 g/cc. Exemplary densities include about1.058 g/cc to about 1.070 g/cc, including about 1.064 g/cc. Such a buoydesign effects a substantially maximum change in displacement of avolume of fractionated suspension below an equilibrium interface andeffects a substantially maximum change in displacement of a volume offractionated suspension above the equilibrium interface relative to theaxial displacement of the buoy resulting in more precisely controllingthe selected fraction isolation. As discussed above, the buoy, accordingto various embodiments, has a maximum cross section at a selectedregion. Positioning a maximum cross section within a selected fractionor area of a sample will maximum displacement of the sample relative tothe buoy do to the maximum cross section of the buoy. In other words, bypositioning the biggest portion of the buoy within a selected sample thebiggest portion of the sample is displaced because of the displacementof the buoy.

Particulates are concentrated using a guide surface 42, 90, 138, 152,162 of the buoy that is inclined to an accumulation position near aperimeter of the buoy. The guide surface can be inclined relative to thebuoy sidewall substantially throughout a length of the guide surface.The guide surface can be defined by or positioned near the top wall ofthe buoy.

The particulates are conveyed along the guide surface of the buoy to acollection space. The particulates can be conveyed along a substantiallyplanar path to the collection space. According to various embodiments,however, the guide surface can also include multiple angles 42, 44 and152, 154 and/or contours 96, 98. The particulates can be selected fromthe group consisting of platelets, stromal cells, white blood cells, orthe like.

A desired fraction is withdrawn from the collection space through anaccess port. In some embodiments, the desired fraction can be withdrawnfrom the collection space by tipping the separation container andpouring the desired fraction out through an access port or out throughthe container top. This is especially true when only the buoy 30′, 30″,30′″ is present (FIGS. 14, 15, and 16).

In some embodiments, the method of isolating a fraction can furthercomprise isolating an isolated fraction in a collection compartmentbetween the guide surface of the buoy 30, 80, 120, 140, 160, 180, 190and an isolator 32, 82, 122, 142, 162, 182, 192 coupled to the buoy andwithdrawing the isolated fraction through a withdraw port through theisolator.

The buoy suspension fractionation system can be used in a method ofisolating and re-suspending particulates for withdrawal. The methodbegins by filling a separation container through an access port with asuspension. The separation container has a buoy with a tuned density andthe suspension can contact the buoy.

The separation container can be centrifuged to cause the suspension toseparate into fractions of varying densities. Centrifugation can occurfor a period that is appropriate for the suspension, such as about fiveto about thirty minutes.

The buoy in the separation container is allowed to reach equilibriumwithin the fluid between two or more fractions. Typically the buoy movesfrom the separation container bottom to equilibrium within thefractions. In some embodiments, particulates can be concentrated using aguide surface of the buoy. The guide surface can be inclined to anaccumulation position 44, 92 near a buoy perimeter location. Accordingto various embodiments, the guide surface can be inclined relative to abuoy sidewall substantially throughout the length of the guide surface.The particulates can be conveyed along the guide surface of the buoy toa collection port. The particulates can be platelets, stromal cells,white blood cells, or the like.

A fraction is isolated in a collection compartment between the guidesurface of the buoy and an isolator coupled to the buoy. In someembodiments, there can be a fraction 308 located above the isolator thatcan be withdrawn prior to withdrawing a first increment of the secondfraction 310. In other embodiments, the collection vent tube 203 caneliminate the need to withdraw the fraction 308 located above theisolator prior to withdrawing the first increment of the second fraction310.

Particulates within the isolated fraction can be re-suspended within thecollection compartment by moving an agitator 130, 316 (FIGS. 11A and 19)in the separation container 12 to agitate the isolated fraction tocreate a more uniform particulate distribution within the isolatedfraction. In some embodiments, the agitator is an air bubble 316 that iscreated by withdrawing the first increment of the isolated fraction 310from a collection compartment allowing air to enter the collectioncompartment through the collection vent 58. In other embodiments, theagitator 130 is a mechanical agitator placed in the collectioncompartment.

The re-suspended isolated fraction can be withdrawn from the collectioncompartment.

For illustration and for efficiency of use of the system, the variouscomponents can be included in a kit 320, illustrated in FIG. 20. The kit320 includes the fractionation system 10 and a counterweight container322 if required for centrifuge balance. The kit 320 can also includevarious syringes 302, 312, and 314 for extraction and application of thefractions and samples. The kit 320 can also include bandages 3226, tape330, a tourniquet 328, and various additive materials. The kit 320 caninclude a container 332 for transport and sterilization.

Thus, embodiments of a buoy suspension fractionation system aredisclosed. One skilled in the art will appreciate that the teachings canbe practiced with embodiments other than those disclosed. The disclosedembodiments are presented for purposes of illustration and notlimitation, and the invention is only limited by the claims that follow.

1. A buoy suspension fractionation system, comprising: a separationcontainer and an access port through at least a portion of theseparation container, the separation container is enclosed by acontainer wall, a container bottom, and a container top; and, a buoy inthe separation container, the buoy having a tuned density and configuredto reach an equilibrium position in a suspension material, the buoycomprising a buoy upper surface and a buoy sidewall defining a height, atransverse dimension through a center portion of the buoy, and aperimeter, the buoy further comprising; a guide surface on the buoyupper surface, the guide surface having a collection space above thebuoy upper surface and the guide surface being inclined to anaccumulation position near the buoy perimeter; and a collection portextending from the guide surface nearer the buoy perimeter than thecenter portion of the buoy and communicating with the access port andcommunicating with the collection space above the buoy upper surface. 2.The buoy suspension fractionation system as in claim 1 wherein the buoyincludes a maximum transverse dimension configured to be positioned nearan equilibrium interface between two separated fractions of thesuspension and substantially defining a buoy maximum cross-sectionalarea.
 3. The buoy suspension fractionation system as in claim 2 whereinmovement of the buoy near an equilibrium interface results insubstantially maximum change in displacement of a fractionatedsuspension below the equilibrium interface and substantially maximumchange in displacement of a fractionated suspension above theequilibrium interface relative to change in axial orientation of thebuoy relative to the interface.
 4. The buoy suspension fractionationsystem as in claim 1 wherein the guide surface is inclined relative tothe buoy sidewall for a distance of more than about two-thirds the buoytransverse dimension.
 5. The buoy suspension fractionation system as inclaim 4 wherein the guide surface is inclined relative to the buoysidewall substantially throughout an entire distance of the guidesurface.
 6. The buoy suspension fractionation system as in claim 1wherein the guide surface is inclined in relation to the buoy sidewallat an average angle of about 10 degrees to about 70 degrees.
 7. The buoysuspension fractionation system as in claim 1 wherein the guide surfaceis substantially planar for the entire transverse dimension of the buoy.8. The buoy suspension fractionation system as in claim 1 wherein theguide surface is inclined substantially outwardly toward the buoyperimeter.
 9. The buoy suspension fractionation system as in claim 8,wherein the guide surface extends from a first edge of the buoyperimeter along the transverse dimension and substantially through acenter of the buoy to a second edge of the buoy perimeter different fromthe first edge.
 10. The buoy suspension fractionation system as in claim1 further comprising, an isolator coupled to the buoy, the isolatorcomprising a spacer to position the isolator apart from the buoy tocreate a collection area within the collection space.
 11. The buoysuspension fractionation system as in claim 10 further comprising, awithdraw port carried on the isolator communicating with the access portand the collection area.
 12. The buoy suspension fractionation system asin claim 10 wherein the isolator has an isolator volume and the buoy hasa buoy volume; wherein a first combination of both of the isolatorvolume and the buoy volume below an equilibrium interface is operable tobe greater than a second combination of both of the isolator volume andthe buoy volume above the equilibrium interface.
 13. The buoy suspensionfractionation system as in claim 10 wherein the isolator is configuredto move from an open position apart from the buoy to a closed positionnear the buoy.
 14. The buoy suspension fractionation system as in claim10 wherein the isolator has an isolator volume and the buoy has a buoyvolume; wherein the isolator volume is substantially less than the buoyvolume.
 15. The buoy suspension fractionation system as in claim 10wherein the isolator has collection openings.
 16. The buoy suspensionfractionation system as in claim 10 further comprising, a collectionvent passage through the isolator to equalize pressure when fluid iswithdrawn from the collection area.
 17. The buoy suspensionfractionation system as in claim 16 further comprising, a collectionvalve communicating with the collection vent.
 18. The buoy suspensionfractionation system as in claim 16 further comprising, a collectionvent tube communicating with the collection vent and air above where asuspension would be located in a separation container.
 19. The buoysuspension fractionation system as in claim 10 further comprising, amechanical agitator carried in the collection area.
 20. The buoysuspension fractionation system as in claim 1 wherein the collectionport is formed in a member that extends from the guide surface and islocated near the buoy perimeter and is in direct fluid communicationwith the collection space at the buoy perimeter.
 21. The buoy suspensionfractionation system as in claim 20 further comprising, a precisioncollection region interposed between the guide surface and the buoyperimeter, wherein the precision collection region includes an inclinegreater than the guide surface that extends towards the buoy perimeter;wherein the collection port is in direct fluid communication with theprecision collection region.
 22. The buoy suspension fractionationsystem as in claim 1 wherein when the separation container is at rest,the buoy perimeter and the container wall form an interference fitoperable to hold the buoy at a position in the separation container andwhen the separation container is centrifuged a clearance is definedbetween the buoy perimeter and the container wall allowing the buoy tomove within the separation container and at least a portion of thesuspension material to pass between the buoy perimeter and the containerwall.
 23. The buoy suspension fractionation system as in claim 1 whereinthe separation container is a separation tube.
 24. A buoy for asuspension fractionation system, comprising: a buoy having a tuneddensity and configured to reach an equilibrium point in a suspension,the buoy comprising a buoy upper surface, a buoy sidewall defining aheight, a buoy transverse dimension, and a buoy perimeter, the buoyfurther comprising; a collection port communicating with a collectionspace above the buoy upper surface; a guide surface on the buoy uppersurface, the guide surface being substantially planar and inclined froma first point at an edge of the buoy for a distance of more than twothirds the buoy transverse dimension towards a second point at the edgeof the buoy; an isolator coupled to the buoy, the isolator comprising aspacer to position the isolator apart from the buoy to create acollection compartment in the collection space between the isolator andthe buoy upper surface; and a withdraw port carried on the isolatorcommunicating with the collection port.
 25. The buoy suspensionfractionation system as in claim 24 wherein the buoy transversedimension includes a cross-sectional area of the buoy configured near anequilibrium interface to be substantially a maximum cross-sectional areaof the buoy.
 26. The buoy suspension fractionation system as in claim 25wherein movement of the buoy near an equilibrium interface is operableto result in substantially maximum change in displacement of a fractionbelow the equilibrium interface and substantially maximum change indisplacement of a fraction above the equilibrium interface relative toan axis of buoy relative to the equilibrium interface.
 27. The buoysystem as in claim 24 further comprising, a collection vent on theisolator operable to equalize pressure when fluid is withdrawn from thecollection space.
 28. The buoy system as in claim 24 further comprising,a collection valve communicating with the collection port.
 29. The buoysystem as in claim 28 further comprising, a collection vent tubecommunicating with the collection vent above where a suspension would belocated in a separation container.
 30. The buoy system as in claim 24wherein the collection port is located near the buoy perimeter and is indirect fluid communication with a volume at the second edge.
 31. Thebuoy system as in claim 24 wherein the isolator has collection openings.32. The buoy system as in claim 30 further comprising, a precisioncollection region interposed between the guide surface and the buoyperimeter at the second edge, wherein the precision collection regionincludes an angle of incline towards the second edge that is greaterthan a majority of an angle of the incline of the guide surface.
 33. Thebuoy system as in claim 24 wherein an isolator volume is substantiallyless than a buoy volume.
 34. The buoy system as in claim 24 wherein afirst isolator volume and a first buoy volume combined below anequilibrium interface is operable to be greater than a second isolatorvolume and a second buoy volume combined above the equilibriuminterface.
 35. A buoy suspension fractionation system, comprising: aseparation container and an access port through at least a portion ofthe separation container, the separation container is enclosed by acontainer wall, a container bottom, and a container top; and, a buoy inthe separation container, the buoy having a tuned density and configuredto reach an equilibrium position in a suspension, the buoy comprising abuoy upper surface and a buoy sidewall defining a height, a transversedimension, and a perimeter, the buoy further comprising; a guide surfaceon the buoy upper surface, the guide surface having a collection spaceabove the buoy upper surface and the guide surface being inclined to anaccumulation position near the buoy perimeter; an isolator coupled tothe buoy, the isolator comprising a spacer to position the isolatorapart from the buoy to create a collection area in a space between thebuoy and the isolator; a collection vent passage through the isolator toequalize pressure when fluid is withdrawn from the collection area; anda collection valve communicating with the collection vent.
 36. The buoysuspension fractionation system as in claim 35 wherein the guide surfaceis inclined from one point on the buoy sidewall to a second point on thebuoy sidewall though a center portion of the buoy for a distance of morethan about two-thirds the buoy transverse dimension.
 37. The buoysuspension fractionation system as in claim 35 wherein the guide surfaceis inclined relative to the buoy sidewall in one direction substantiallythroughout a length of the guide surface.
 38. The buoy suspensionfractionation system as in claim 35 wherein the guide surface issubstantially planar.
 39. The buoy suspension fractionation system as inclaim 35, wherein the guide surface extends from a first edge of thebuoy perimeter along the transverse dimension and substantially througha center of the buoy to a second edge of the buoy perimeter; wherein theguide surface defines at least two thirds a width of the buoy.
 40. Thebuoy suspension fractionation system as in claim 35, further comprisinga collection port communicating with the access port and communicatingwith the collection space above the buoy upper surface.
 41. The buoysuspension fractionation system as in claim 35 further comprising, awithdraw port carried on the isolator communicating with the access portand the collection area.
 42. The buoy suspension fractionation system asin claim 35 wherein the isolator has an isolator volume and the buoy hasa buoy volume; wherein the isolator volume is substantially less thanthe buoy volume.
 43. The buoy suspension fractionation system as inclaim 40 wherein the collection port is in communication with a volumeat the buoy perimeter.
 44. The buoy suspension fractionation system asin claim 43 further comprising, a precision collection region interposedbetween the guide surface and the buoy perimeter in the volume at thebuoy perimeter.
 45. A buoy suspension fractionation system, comprising:a separation container and an access port through at least a portion ofthe separation container, the separation container is enclosed by acontainer wall, a container bottom, and a container top; and, a buoy inthe separation container, the buoy having a tuned density and configuredto reach an equilibrium position in a suspension, the buoy comprising abuoy upper surface and a buoy sidewall defining a height, a transversedimension, and a perimeter, the buoy further comprising; a guide surfaceon the buoy upper surface, the guide surface having a collection spaceabove the buoy upper surface and the guide surface being inclined to anaccumulation position near the buoy perimeter; an isolator coupled tothe buoy, the isolator comprising a spacer to position the isolatorapart from the buoy to create a collection area in a space between thebuoy and the isolator; a collection vent passage through the isolator toequalize pressure when fluid is withdrawn from the collection area; anda collection vent tube communicating with the collection vent and airabove where a suspension would be located in a separation container. 46.The buoy suspension fractionation system as in claim 45 wherein the buoyincludes a maximum transverse dimension configured to be positioned nearan equilibrium interface between two separated fractions of thesuspension and substantially defining a buoy maximum cross-sectionalarea.
 47. The buoy suspension fractionation system as in claim 45wherein the guide surface is inclined relative to the buoy sidewall inone direction substantially throughout a length of the guide surface.48. The buoy suspension fractionation system as in claim 45 wherein theguide surface is substantially planar.
 49. The buoy suspensionfractionation system as in claim 45, wherein the guide surface extendsfrom a first edge of the buoy perimeter along the transverse dimensionand substantially through a center of the buoy to a second edge of thebuoy perimeter; wherein the guide surface defines at least two thirds awidth of the buoy.
 50. The buoy suspension fractionation system as inclaim 45, further comprising a collection port communicating with theaccess port and communicating with the collection space above the buoyupper surface.
 51. The buoy suspension fractionation system as in claim45 further comprising, a withdraw port carried on the isolatorcommunicating with the access port and the collection area.
 52. The buoysuspension fractionation system as in claim 45 further comprising, aprecision collection region interposed between the guide surface and thebuoy perimeter in the volume at the buoy perimeter.
 53. The buoysuspension fractionation system as in claim 24, wherein the buoytransverse dimension passes through a center of the buoy.